CN114480201A - Vibrio natriegens capable of strongly degrading enteromorpha protein and application of vibrio natriegens in preparation of organic fertilizer - Google Patents
Vibrio natriegens capable of strongly degrading enteromorpha protein and application of vibrio natriegens in preparation of organic fertilizer Download PDFInfo
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Abstract
The invention discloses a vibrio natriegens capable of strongly degrading enteromorpha protein and application thereof in preparation of an organic fertilizer. The invention collects a water sample separation strain containing rotten enteromorpha from a sea water bathing field in yellow island region of Qingdao city in Shandong province, and screens out a strain Vibrio natriegens (Vibrio natriegens) HT-7 with an enteromorpha degradation effect through a degradation experiment, wherein the preservation number of the strain is CGMCC NO. 23444. The bacterial strain can efficiently degrade enteromorpha crude protein, can fully degrade enteromorpha to prepare organic fertilizer, and the prepared organic fertilizer can promote the growth of various plants in different families and has the broad-spectrum growth promotion property. Meanwhile, the strain microbial inoculum is low in production cost, simple in equipment, mild in reaction, suitable for large-scale industrial production, and good in application potential in the aspect of effective treatment of enteromorpha.
Description
Technical Field
The invention relates to vibrio natriegens capable of strongly degrading enteromorpha protein and application of vibrio natriegens in preparation of an organic fertilizer, and belongs to the technical field of microorganisms.
Background
Enteromorpha (Enteromorpha prolifera) is a large green algae commonly called as Enteromorpha and sea moss, and is a plant of Ulvaceae of Ulvales of the green algae Menispermales. In recent years, the phenomenon of seawater eutrophication caused by mass propagation of algae such as enteromorpha has become an important expression of marine environmental pollution, and a large amount of algae salvaged in the treatment process is difficult to dispose. At present, temporary stockpiling and landfill are common enteromorpha fast disposal modes, but secondary pollution is generated in the process, and mainly comprises the influence of leachate and malodorous gas on the environment. Therefore, how to realize the harmless comprehensive utilization of the enteromorpha becomes a current research hotspot.
Because the enteromorpha prolifera contains rich crude protein, polysaccharide and crude fiber, the enteromorpha prolifera can be used as effective raw materials for various resource utilization, such as biological compost, including production of liquid fertilizers and solid fertilizers, feed production, food processing and energy utilization and the like. However, the high-efficiency degradation of the enteromorpha becomes an important factor for restricting the recycling of the enteromorpha. At present, the enteromorpha degrading method mainly comprises a physical method and a chemical method, but the two methods have certain disadvantages, such as acid degradation in the chemical method, poor substrate specificity due to violent action conditions, and the sulfate group of the sulfated polysaccharide is easy to hydrolyze and fall off in the degrading process, so that the difficulty of separation and purification is increased, and the cost is high; physical degradation modes like natural fermentation are inefficient and inflexible. Therefore, the method becomes an effective way for solving the problems of low efficiency, high cost and the like in the enteromorpha degradation process by screening the microbial degradation bacteria.
But few researches on the enteromorpha high-efficiency degrading bacteria are carried out at present. The patent CN 108239613B discloses a paenibacillus foraging, a culture medium thereof and application thereof in preparing enteromorpha polysaccharide degrading enzyme, wherein the microorganism preservation number is CGMCC NO.12912, and the enteromorpha polysaccharide degrading enzyme produced by the strain has good enzyme activity, but the strain is not applied to industrial production. The patent CN 103695344A discloses a bacterial strain of Bacillus subtilis and a screening method and application thereof, wherein the microorganism preservation number of the bacterial strain is CGMCC NO.8269, the bacterial strain has the capability of producing cellulase, can effectively degrade cellulose substances in enteromorpha into reducing sugar, and is also lack of an enteromorpha comprehensive utilization process. Therefore, at present, the types of the strains are few in the utilization aspect of the enteromorpha high-efficiency degrading bacteria, reports that the enteromorpha is degraded by vibrio natriegens are not shown, and reports that the enteromorpha is subjected to resource utilization on the organic fertilizer produced by degrading the enteromorpha by the vibrio natriegens are not shown.
Disclosure of Invention
Aiming at the problems, the invention provides a Vibrio natriegens strain capable of strongly degrading enteromorpha protein and an application thereof, firstly, a Vibrio natriegens strain HT-7 with the capability of remarkably degrading crude enteromorpha protein is screened, and the enteromorpha protein degrading organic fertilizer protoplasm prepared by degrading enteromorpha by adopting the strain can promote the growth of various plants in different families and has the growth promoting broad spectrum.
The strain HT-7 provided by the invention is separated from a water sample containing rotten enteromorpha collected from a sea water bath field in yellow island region of Qingdao city in Shandong province, is a high-efficiency strain which is screened by a degradation experiment and has a degradation effect on enteromorpha, and is identified as Vibrio natriegens (Vibrio natriegens) by combining morphological and gene sequence determination results. The strain has been preserved in China general microbiological culture Collection center (CGMCC) at 18 days 9 months 2021, the preservation number is CGMCC NO.23444, the preservation unit address, No. 3 of Xilu 1 north Chen in the rising area of Beijing. Experiments prove that: the strain has the capacity of remarkably degrading enteromorpha crude protein, and has wide prospects in the fields of enteromorpha treatment and organic fertilizer preparation.
The above-mentioned strain of Vibrio natriegens HT-7 is characterized in that: the culture medium is irregular and round on an NA culture medium, is light yellow, has a smooth and moist surface and regular edges, and has a diameter of 1-2 mm.
The invention also provides a fermentation method of the vibrio natriegens HT-7, which is characterized in that seed liquid of the vibrio natriegens HT-7 is inoculated into a fermentation medium according to the inoculation amount of 1-2%, and the seed liquid is fermented and cultured for 24 +/-4 h at the constant temperature of 20 +/-2 ℃ to obtain fermentation liquid (fermentation microbial inoculum) of the vibrio natriegens, wherein the fermentation medium comprises the following components in percentage by weight: 1.5-2% of glucose, 1.8-2% of starch, 1.3-1.5% of yeast extract powder, 0.3-0.5% of fish peptone, 0.04-0.06% of magnesium sulfate, 0.04-0.06% of potassium dihydrogen phosphate, 0.5-0.8% of calcium carbonate and 7.2 +/-0.1% of pH value.
The invention also provides an application of the vibrio natriegens HT-7 in degrading enteromorpha to prepare an organic fertilizer, wherein the degraded enteromorpha comprises crude protein, polysaccharide, crude fat and dry matter, especially crude protein, of the degraded enteromorpha.
The invention also provides a method for preparing an organic fertilizer by degrading enteromorpha through vibrio natriegens HT-7, which is characterized by comprising the following steps of:
pretreatment of enteromorpha prolifera: soaking and rinsing the collected enteromorpha prolifera in fresh water to remove salt; then fully crushing;
fully mixing the crushed enteromorpha prolifera with water (the mass ratio is 1.5-2.5: 1), and then feeding into a fermentation tank;
inoculating the fermentation liquor requiring vibrio natriens HT-7 according to the inoculation amount of 1.0-5.0% (mass ratio), maintaining the temperature of the tank at 20 +/-2 ℃ after inoculation for 5-6 days, taking out the tank after fermentation, and filtering. The liquid is the original pulp of the enteromorpha prolifera degradation protein organic fertilizer, and can be matched with other medium and trace elements to produce various organic fertilizers; the residual solid can be used as raw material for producing livestock and poultry feed.
Furthermore, aeration is not carried out in the process of inoculating the fermentation liquor of the vibrio natriegens HT-7; fully stirring and fermenting after inoculation is finished, only ventilating a small amount (the ventilation amount is 0.2:1M.M/V), and fermenting for 3 days; then, a large amount of air (the air flow is 1:1M.M/V) is aerated, and the mixture is continuously and fully stirred and fermented for 2-3 days.
The invention has the technical effects that:
1. after the Enteromorpha prolifera is treated by the fermentation inoculum of the strain, the high efficiency of the strain is shown in degrading Enteromorpha prolifera crude protein, the crude protein is reduced by 12.3 percent, and the contents of dry substances, polysaccharides and crude fat are also reduced to different degrees.
2. After the vibrio natriegens HT-7 fermentation microbial inoculum and the fully crushed enteromorpha are effectively mixed, the enteromorpha is fully degraded after being fermented for 5d, the produced liquid can be used as a raw material for producing organic water-soluble fertilizers, and the prepared organic fertilizers can promote the growth of various plants in different families and have broad growth promotion spectrum.
3. The vibrio natriegens HT-7 is easy to ferment and produce, low in production cost, simple in equipment, mild in reaction, suitable for large-scale industrial production and good in application potential in the aspect of effective disposal of enteromorpha.
Drawings
FIG. 1 is a colony morphology of Vibrio natriegens HT-7;
FIG. 2 is the appearance state of Enteromorpha after treating Vibrio natrii HT-7 in a test tube for different days;
FIG. 3 is the appearance state of Enteromorpha prolifera 9d treated by Vibrio natrii HT-7 microbial inoculum in a sealed bag.
Detailed Description
The technical solutions in the embodiments of the present invention will be clearly and completely described below with reference to the contents in the embodiments of the present invention, and it is obvious that the described embodiments are only a part of the embodiments of the present invention, and not all of the embodiments. All other embodiments, which can be derived by a person skilled in the art from the embodiments given herein without making any creative effort, shall fall within the protection scope of the present invention.
Example 1: isolation and characterization of strains
1. Isolation of Strain HT-7
Separating on NA culture medium by dilution coating method, wherein the sample is water sample containing rotten Enteromorpha prolifera collected from sea water bathing ground in yellow island region of Qingdao, Shandong province. The specific separation method is as follows:
(1) and (3) sterilization: sterilizing the culture dish, the gun head, the sterile water and other experimental articles under 0.1MPa and 121 ℃ for 30min for later use;
(2) bacterial sample dilution: mixing the water sample and the smashed and rotten enteromorpha according to a ratio of 9:1, placing the mixture in an aseptic triangular flask containing glass beads, oscillating the mixture for 30min under the condition of 120r/min, and filtering the mixture by four layers of aseptic gauze to obtain a bacterial sample; 6 test tubes containing 9mL of sterile water are sequentially numbered 10-1～10-6(ii) a Sucking 1mL of the bacteria sample from the triangular flask and putting the bacteria sample into a sample bottle marked with 10-1In a test tube, i.e. 10-1Diluting; and again indicated with 10-1Taking 1mL of the diluted bacterial solution, and putting the diluted bacterial solution into a container marked with 10-2In a test tube, i.e. 10-2Diluting; sequentially prepared by the same method to obtain 10-3、10-4、10-5、10-6A diluted bacterial sample;
(4) preparation of the Medium
Isolation medium, i.e. NA medium: 10g of peptone, 3g of beef powder, 5g of NaCl, 15g of agar and 1000mL of water, wherein the pH value is 7.3 +/-0.1;
purifying the culture medium and separating the culture medium;
(5) coating: diluting the bacterial sample 10-3、10-4、10-5、10-6Respectively coating on a separation culture medium flat plate, each gradient 3 dish, and marking on the bottom of the dish;
(6) culturing: after coating, carrying out inverted culture at a constant temperature of 20 ℃ for about 3 days, and observing;
(8) strain purification: picking single colony with inoculating hook, streaking on plate containing purified culture medium, and reverse culturing at 20 deg.C for 3d to pick single colony.
As a result, 8 strains of bacteria were isolated in total and numbered HT-1 to HT-8, respectively.
2. Strain HT-7 screening
Culturing the strains with different numbers obtained by the separation for 24 hours by using a shake flask culture method (see example 2) to respectively prepare shake flask culture solutions of the strains; weighing 5g of fresh enteromorpha and placing the fresh enteromorpha in a test tube to keep the consistency of compactness; adding 1mL of shake flask microbial inoculum of different strains into each test tube, plugging a plug and providing an anaerobic environment; and (5) observing at 20 ℃ after 5 d.
The result shows that the enteromorpha inoculated with the HT-7 microbial inoculum becomes black after being treated for 5 days; the color of the enteromorpha inoculated with HT-1 and HT-3 bactericides is changed from green to light. The results preliminarily show that HT-1, HT-3 and HT-7 have the function of degrading enteromorpha, and the effect of HT-7 is superior to that of other strains. Next, we performed subsequent studies with HT-7 as the subject.
TABLE 1 summary of degradation of Enteromorpha by different strains
The criteria for judging the activity level are: the enteromorpha is black-starting grade of +++, the enteromorpha is brown and hydrated and is of +++, the enteromorpha is green-lightening grade of ++, and the enteromorpha is non-discoloring grade of- ".
3. Morphological and molecular biological identification of strain HT-7
(1) Morphological characteristics: the strain is irregularly round on an NA culture medium, is light yellow, has a smooth and moist surface and regular edges, and has a diameter of 1-2 mm (refer to FIG. 1).
(2) Molecular biological Properties
The sequence determination result of the 16s rDNA gene of the strain is as follows (SEQ-1):
AGTAATGGTTAAACTACCTACTTCTTTTGCAGCCCACTCCCATGGTGTGACGGGCGG TGTGTACAAGGCCCGGGAACGTATTCACCGTGGCATTCTGATCCACGATTACTAGCGATT CCGACTTCATGGAGTCGAGTTGCAGACTCCAATCCGGACTACGACGCACTTTTTGGGAT TCGCTCACTTTCGCAAGTTGGCCGCCCTCTGTATGCGCCATTGTAGCACGTGTGTAGCCC TACTCGTAAGGGCCATGATGACTTGACGTCGTCCCCACCTTCCTCCGGTTTATCACCGGC AGTCTCCCTGGAGTTCCCGACATTACTCGCTGGCAAACAAGGATAAGGGTTGCGCTCGT TGCGGGACTTAACCCAACATTTCACAACACGAGCTGACGACAGCCATGCAGCACCTGTC TCAGAGTTCCCGAAGGCACCAATCCATCTCTGGAAAGTTCTCTGGATGTCAAGAGTAGG TAAGGTTCTTCGCGTTGCATCGAATTAAACCACATGCTCCACCGCTTGTGCGGGCCCCCG TCAATTCATTTGAGTTTTAATCTTGCGACCGTACTCCCCAGGCGGTCTACTTAACGCGTTA GCTCCGAAAGCCACGGCTCAAGGCCACAACCTCCAAGTAGACATCGTTTACGGCGTGG ACTACCAGGGTATCTAATCCTGTTTGCTCCCCACGCTTTCGCATCTGAGTGTCAGTATCTG TCCAGGGGGCCGCCGTTCGCCACCGGTATTCCTTCAGATCTCTACGCATTTCACCGCTAC ACCTGAAATTCTACCCCCCTCTACAGTACTCTAGTCTGCCAGTTTCAAATGCTATTCCGA GGTTGAGCCCCGGGCTTTCACATCTGACTTAACAAACCACCTGCATGCGCTTTACGCCC AGTAATTCCGATTAACGCTCGCACCCTCCGTATTACCGCGGCTGCTGGCACGGAGTTAGC CGGTGCTTCTTCTGTCGCTAACGTCAAATAAAGCATCTATTAAACCCAAAACCTTCCTCA CGACTGAAAGTGCTTTACAACCCGAAGGCCTTCTTCACACACGCGGCATGGCTGCATCA GGCTTGCGCCCATTGTGCAATATTCCCCACTGCTGCCTCCCGTAGGAGTCTGGACCGTGT CTCAGTTCCAGTGTGGCTGATCATCCTCTCAGACCAGCTAGGGATCGTCGCCTTGGTGA GCCCTTACCTCACCAACTAGCTAATCCCACCTAGGCATATCCTGACGCGAGAGGCCCGA AGGTCCCCCTCTTTGGCCCGTAGGCATCATGCGGTATTAGCCATCGTTTCCAATGGTTATC CCCCACATCAGGGCAATTTCCTAGGCATTACTCACCCGTCCGCCGCTCGACGCCGTTAAC GTCCCCCGAAGGTTCAGTTAACTCGTTTCCGCTCGACTGCATGGTAGTCAGCGCCAGC
the strain HT-7 is morphologically and molecularly identified as Vibrio natriegens (Vibrio natriegens), which has been deposited in the general microbiological center of China Committee for culture Collection of microorganisms with the date of deposition: 18 months 9 in 2021, with the preservation number of CGMCC NO. 23444.
Example 2: preparation of Vibrio natriegens HT-7 fermentation microbial inoculum
(1) Dipping a small amount of purified strains by using an inoculating loop into a 50mL triangular flask containing 10mL of seed liquid culture medium, and culturing at constant temperature of 120r/min and 20 ℃ for 24h to obtain seed liquid;
seed liquid culture medium, i.e. NB-containing medium: 10g of peptone, 3g of beef powder, 5g of NaCl and 1000mL of water, wherein the pH value is 7.3 +/-0.1;
(2) inoculating the seed solution prepared in the step (1) into a fermentation culture medium, wherein the inoculation amount is 1.5%, fermenting and culturing at constant temperature of 20 ℃ for 24h to obtain fermentation liquor (shake flask microbial inoculum) of Vibrio natrii HT-7, and the viable count of the fermentation liquor is 2-5 multiplied by 1010cfu/g. The shake flask microbial inoculum of each strain can be obtained by fermenting each strain in a shake flask.
Fermentation medium (mass ratio): 2% of glucose, 1.8% of starch, 1.5% of yeast extract powder, 0.4% of fish peptone, 0.05% of magnesium sulfate, 0.05% of monopotassium phosphate, 0.6% of calcium carbonate, 1L of water and pH7.2 +/-0.1.
Example 3: research on degradation function of Vibrio natriegens HT-7
(1) Functional verification of vibrio natriegens HT-7 degradation enteromorpha
See example 2 for making a Vibrio natriegens HT-7 fermentation inoculum; weighing 5g of fresh enteromorpha and placing the fresh enteromorpha in a test tube to keep the consistency of compactness; adding 1mL of vibrio natriegens HT-7 microbial inoculum into a test tube, and adding 1mL of fermentation medium into a control group; each treatment was repeated 3 times; the plugs are plugged uniformly to provide an anaerobic environment; and (5) observing at 3d, 5d, 7d and 9d respectively under the condition of 20 ℃, and recording the appearance state of the enteromorpha.
As a result, compared with the control, after the Vibrio natriegens HT-7 microbial inoculum is applied for 3d, the color of the enteromorpha is changed from green to light; after the enteromorpha is treated for 5 days, the browning color of the enteromorpha is deepened, and hydration is started; after the treatment for 7d, the enteromorpha starts to blacken; after 9d of treatment, all enteromorpha was blackened (see fig. 2). In conclusion, the research results preliminarily prove that HT-7 obtained by screening has excellent enteromorpha degrading capability, and the results are consistent with the screening results; meanwhile, the degradation function of the Vibrio natriegens HT-7 microbial inoculum is stable.
(2) Vibrio natriegens HT-7 degradability assay
Preparing a fermentation inoculum of Vibrio natriegens HT-7 according to example 2; weighing 1Kg of fresh enteromorpha and placing in a sealed bag; adding 200mL of Vibrio natriegens HT-7 microbial inoculum into the sealed bag, and adding 200mL of fermentation medium into the control group; each treatment was repeated 3 times; sealing to provide an anaerobic environment; and (5) observing at 9d under the condition of 20 ℃, and recording the appearance state of the enteromorpha prolifera.
The dry matter content, polysaccharide content, crude protein content and crude fat content of enteromorpha before and after treatment were respectively detected by a drying and weighing method, a phenol-sulfuric acid method, a Kjeldahl semi-micro nitrogen determination method and a Soxhlet extraction method, and the content change of four indexes was counted, as shown in Table 2.
The result analysis shows that after the Enteromorpha prolifera is treated by the Vibrio natriensis HT-7 microbial inoculum for 9 days, the Enteromorpha prolifera is partially blackened and hydrated (refer to figure 3); the color of part of the enteromorpha of the control group begins to become lighter; this preliminarily shows that Vibrio natriegens HT-7 has strong ability to degrade Enteromorpha. Through content detection of dry matter, polysaccharide, crude protein and fat of enteromorpha before and after treatment, after treatment by a vibrio natriensis HT-7 microbial inoculum, the dry matter is reduced by 25.5%, the polysaccharide matter is reduced by 9.7%, the crude protein is reduced by 12.3%, the crude fat is reduced by 3.1% (see table 2), and the capacity of degrading the crude protein is remarkable.
TABLE 2 variation of the content of relevant components of Enteromorpha after Vibrio natriegens HT-7 treatment
Meanwhile, the degradation product obtained after full fermentation and degradation of the enteromorpha is sent to Suzhou Stentde laboratory science and technology limited company for detection, and the total amount of glucose, amino acid and polypeptide and the content of fatty acid are detected. The result shows that after the vibrio natriegens HT-7 is degraded, the main components of the enteromorpha degradation product are micromolecule glucose, amino acid, polypeptide and fatty acid, the contents are 39.6%, 18.5% and 0.1%, respectively, and the crude protein degradation capability of the enteromorpha degradation product is obvious.
TABLE 3 major Components of Enteromorpha degradants after HT-7 treatment of Vibrio natriegens
Example 4: research on effect of enteromorpha prolifera degradation protein organic fertilizer protoplasm
(1) Process route
On the basis of screening Vibrio natriegens HT-7 and function preliminary verification, a process route for treating enteromorpha by HT-7 is researched.
Pretreatment of enteromorpha prolifera: rinsing the collected enteromorpha prolifera by brewing with fresh water, removing salt, and repeating for 3-5 times; fully crushing by a crusher.
② fully mixing the crushed enteromorpha with water (mass ratio is 2:1), putting the mixture into a 10-ton fermentation tank, and canning 80 percent;
inoculating a Vibrio natriegens HT-7 fermentation inoculum (prepared in example 2) according to the inoculation amount of 1.5%, and inoculating the Vibrio natriegens HT-7 fermentation inoculum while feeding; maintaining the rotation speed at 60-80 r/min, and keeping the ventilation at the moment.
Fourthly, after inoculation is finished, the temperature of the tank is maintained at 20 ℃, the fermentation is fully stirred at 100r/min, and only a small amount of air is introduced (the air flow is 0.2:1M.M/V), and the fermentation is carried out for 3 d.
Fifthly, ventilating a large amount (the ventilation amount is 1:1M.M/V) after fermenting for 3d, maintaining the temperature of the tank at 20 ℃, and fully stirring at 120r/min for fermenting for 2 d.
Sixthly, taking out of the tank after fermentation is finished, filtering to obtain liquid, namely the enteromorpha degraded protein organic fertilizer raw stock, and producing various organic fertilizers by matching with other medium and trace elements; the residual solid can be used as raw material for producing livestock and poultry feed.
(2) Effect of enteromorpha prolifera degradation protein organic fertilizer protoplasm on growth of different crops
In order to verify the protoplasm growth promotion performance of the enteromorpha prolifera degradation protein organic fertilizer, materials with certain economic values in different families (cucurbitaceae, cruciferae, leguminosae and solanaceae) are selected to carry out a related growth promotion experiment.
Experimental groups: taking plant materials with consistent growth vigor after transplanting, diluting the produced enteromorpha degraded protein organic fertilizer raw stock by 500 times, and carrying out root irrigation experiments, wherein the inoculation amount is 75 mL/plant; control group: each plant was inoculated with 75mL of water; water twice a week. And after 15 days of inoculation, comparing the growth vigor of the plants in the control group and the experimental group, and detecting the related growth indexes. The result shows that the original pulp inoculated with the enteromorpha degraded protein organic fertilizer has the growth promoting effect of different degrees on materials of different families, and particularly has more obvious promoting effect on solanaceae crops; wherein the plant height of the pepper is increased by 25 percent, the plant height of the tomato is increased by 22.24 percent, and the leaf area and the dry weight are increased to a certain degree. The enteromorpha degradation protein organic fertilizer raw pulp obtained by utilizing the enteromorpha after HT-7 treatment can be used as a raw material for preparing an organic water-soluble fertilizer, and can be matched with other elements to prepare a novel organic fertilizer.
TABLE 4 influence of original pulp of Enteromorpha prolifera degrading protein organic fertilizer on growth of different crops
SEQUENCE LISTING
<110> Hipposhu Biotech Co., Ltd in Qingdao
<120> vibrio natriegens for strongly degrading enteromorpha protein and application of vibrio natriegens in preparation of organic fertilizer
<130> 0
<160> 1
<170> PatentIn version 3.3
<210> 1
<211> 1428
<212> DNA
<213> 16s rDNA Gene sequence of Vibrio natriegens HT-7
<400> 1
agtaatggtt aaactaccta cttcttttgc agcccactcc catggtgtga cgggcggtgt 60
gtacaaggcc cgggaacgta ttcaccgtgg cattctgatc cacgattact agcgattccg 120
acttcatgga gtcgagttgc agactccaat ccggactacg acgcactttt tgggattcgc 180
tcactttcgc aagttggccg ccctctgtat gcgccattgt agcacgtgtg tagccctact 240
cgtaagggcc atgatgactt gacgtcgtcc ccaccttcct ccggtttatc accggcagtc 300
tccctggagt tcccgacatt actcgctggc aaacaaggat aagggttgcg ctcgttgcgg 360
gacttaaccc aacatttcac aacacgagct gacgacagcc atgcagcacc tgtctcagag 420
ttcccgaagg caccaatcca tctctggaaa gttctctgga tgtcaagagt aggtaaggtt 480
cttcgcgttg catcgaatta aaccacatgc tccaccgctt gtgcgggccc ccgtcaattc 540
atttgagttt taatcttgcg accgtactcc ccaggcggtc tacttaacgc gttagctccg 600
aaagccacgg ctcaaggcca caacctccaa gtagacatcg tttacggcgt ggactaccag 660
ggtatctaat cctgtttgct ccccacgctt tcgcatctga gtgtcagtat ctgtccaggg 720
ggccgccgtt cgccaccggt attccttcag atctctacgc atttcaccgc tacacctgaa 780
attctacccc cctctacagt actctagtct gccagtttca aatgctattc cgaggttgag 840
ccccgggctt tcacatctga cttaacaaac cacctgcatg cgctttacgc ccagtaattc 900
cgattaacgc tcgcaccctc cgtattaccg cggctgctgg cacggagtta gccggtgctt 960
cttctgtcgc taacgtcaaa taaagcatct attaaaccca aaaccttcct cacgactgaa 1020
agtgctttac aacccgaagg ccttcttcac acacgcggca tggctgcatc aggcttgcgc 1080
ccattgtgca atattcccca ctgctgcctc ccgtaggagt ctggaccgtg tctcagttcc 1140
agtgtggctg atcatcctct cagaccagct agggatcgtc gccttggtga gcccttacct 1200
caccaactag ctaatcccac ctaggcatat cctgacgcga gaggcccgaa ggtccccctc 1260
tttggcccgt aggcatcatg cggtattagc catcgtttcc aatggttatc ccccacatca 1320
gggcaatttc ctaggcatta ctcacccgtc cgccgctcga cgccgttaac gtcccccgaa 1380
ggttcagtta actcgtttcc gctcgactgc atggtagtca gcgccagc 1428
Claims (10)
1. Vibrio natriegens (Vibrio natriegens) HT-7 capable of strongly degrading enteromorpha protein is disclosed, wherein the preservation number of the strain is CGMCC NO. 23444.
2. The application of Vibrio natriegens HT-7 in the preparation of organic fertilizer by degrading Enteromorpha prolifera in claim 1.
3. The use according to claim 2, wherein degrading Enteromorpha comprises degrading crude proteins, polysaccharides, crude fats and dry matter of Enteromorpha.
4. The fermentation method of Vibrio natriegens HT-7 according to claim 1, wherein the seed solution of Vibrio natriegens HT-7 is inoculated into the fermentation medium, and the fermentation is performed at a constant temperature of 20 ± 2 ℃ for 24 ± 4h to obtain the fermentation broth.
5. The fermentation process of claim 4, wherein the fermentation medium comprises, by weight: 1.5-2% of glucose, 1.8-2% of starch, 1.3-1.5% of yeast extract powder, 0.3-0.5% of fish peptone, 0.04-0.06% of magnesium sulfate, 0.04-0.06% of potassium dihydrogen phosphate, 0.5-0.8% of calcium carbonate and 7.2 +/-0.1% of pH value.
6. A method for preparing an organic fertilizer by degrading enteromorpha through Vibrio natriegens HT-7 is characterized by comprising the following steps:
pretreatment of enteromorpha prolifera: soaking and rinsing the collected enteromorpha prolifera in fresh water to remove salt; then fully crushing;
secondly, fully mixing the crushed enteromorpha with water, and then feeding the mixture into a fermentation tank;
inoculating the vibrio natriegens HT-7 fermentation liquid prepared by the method of claim 4 or 5 according to the inoculation amount of 1.0-5.0%, maintaining the temperature of the tank at 20 +/-2 ℃ for fermentation for 5-6 days after inoculation, discharging the tank after fermentation is completed, and filtering to obtain the liquid, namely the enteromorpha degraded protein organic fertilizer protoplasm.
7. The method for preparing the organic fertilizer by degrading Enteromorpha prolifera through Vibrio natrii HT-7 as claimed in claim 6, wherein the mass ratio of Enteromorpha prolifera to water is 1.5-2.5: 1.
8. The method for preparing organic fertilizer by degrading enteromorpha with Vibrio natriegens HT-7 as claimed in claim 6, wherein aeration is not performed during the inoculation of Vibrio natriegens HT-7 fermentation broth; fully stirring and fermenting after inoculation is finished, ventilating a small amount of air, and fermenting for 3 days; then ventilating a large amount of air, and continuously and fully stirring and fermenting for 2-3 days.
9. The method for preparing organic fertilizer by degrading Enteromorpha prolifera with Vibrio natriegens HT-7 as claimed in claim 6, wherein the original pulp of the Enteromorpha prolifera degraded protein organic fertilizer prepared in step three is matched with other medium and trace elements to produce various organic fertilizers.
10. The method for preparing organic fertilizer by degrading enteromorpha with Vibrio natriegens HT-7 as claimed in claim 6, wherein the residual solids of step (c) are used as raw material for producing livestock and poultry feed.
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